Our research program is focused on the analysis of the three-dimensional structures of macromolecular assemblies using high resolution electron microscopy. Members of our group work on different, yet, complementary aspects of structural analysis. These include electron crystallographic studies of two-dimensional protein crystals, "single particle" approaches to analyze the structures of protein complexes and determination of the structures of large subcellular assemblies using electron tomography. A significant fraction of our research effort is devoted to developing and implementing novel methods for specimen preparation, high throughput data acquisition and computational analysis. Research Highlights (2001-2002): (a) We have successfully crystallized a bacterial oxalate transporter, and have determined a projection structure at 6 ? resolution, and three-dimensional structure at 6.5 ? resolution. This work provides the first glimpse into the architecture of a very large superfamily of transporter proteins. (b) We have developed methods to introduce a high level of automation in biological electron microscopy. This work has already had a considerable impact on the speed of data collection in our laboratory. NCI has filed a provisional patent application on this technology. (c) We have derived an atomic model for the 11,000 kD pyruvate dehydrogenase complex from B. stearothermophilus. The model was obtained by docking in four individual components of the enzyme whose structures have been determined by X-ray crystallography into a medium resolution electron microscopic model of the entire complex. The results have been very exciting and unexpected. (d) We are developing tools to carry out three-dimensional microscopy of whole cells and sections using newly emerging methods in electron tomography. A variety of projects ranging from imaging bacteria to whole brain tissue are underway. (e) We have demonstrated that CCD detectors can now fully replace photographic film to record electron diffraction patterns to resolution of 2 ?, and record images for structure determination from "single particles" to medium resolution (~ 10 ?).